The differential functional stability of various forms of bovine liver rhodanese

Aird, Bruce A.; Horowitz, Paul M.

doi:10.1016/0167-4838(88)90294-4

Cited by 13 publications

(5 citation statements)

References 30 publications

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“…The presence of thiosulfate during the purification steps [27] was required for bovine liver rhodanese, since the E form of this enzyme was very sensitive to oxidative reactions [28]. In addition, the functional stability of bovine rhodanese in the sulfur‐substituted form (ES) could be increased in the presence of thiosulfate, this latter being an effective scavenger of free radicals in solution [29]. In contrast, the functional stability of A. vinelandii RhdA was not affected by the presence of thiosulfate, and the enzyme appeared more stable compared to the bovine enzyme (data not shown).…”

Section: Resultsmentioning

confidence: 99%

Mutagenic analysis of Thr‐232 in rhodanese from Azotobacter vinelandii highlighted the differences of this prokaryotic enzyme from the known sulfurtransferases

et al. 2000

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Azotobacter vinelandii RhdA uses thiosulfate as the only sulfur donor in vitro, and this apparent selectivity seems to be a unique property among the characterized sulfurtransferases. To investigate the basis of substrate recognition in RhdA, we replaced Thr-232 with either Ala or Lys. Thr-232 was the target of this study since the corresponding Lys-249 in bovine rhodanese has been identified as necessary for catalytic sulfur transfer, and replacement of Lys-249 with Ala fully inactivates bovine rhodanese. Both T232K and T232A mutants of RhdA showed significant increase in thiosulfate-cyanide sulfurtransferase activity, and no detectable activity in the presence of 3-mercaptopyruvate as the sulfur donor substrate. Fluorescence measurements showed that wild-type and mutant RhdAs were overexpressed in the persulfurated form, thus conferring to this enzyme the potential of a persulfide sulfur donor compound. RhdA contains a unique sequence stretch around the catalytic cysteine, and the data here presented suggest a possible divergent physiological function of A. vinelandii sulfurtransferase.z 2000 Federation of European Biochemical Societies.

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Section: Resultsmentioning

confidence: 99%

Mutagenic analysis of Thr‐232 in rhodanese from Azotobacter vinelandii highlighted the differences of this prokaryotic enzyme from the known sulfurtransferases

et al. 2000

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“…To rule out the possibility that NP-mediated islet-cell lysis is caused by cyanide, the latter was converted to untoxic thiocyanate by bovine liver rhodanese (thiocyanate :cyanide-sulphurtransferase, EC 2.8.1.1.) [25] which has been shown to be active at 37 ~ for at least 15 h in the presence of thiosulphate [26]. Rhodanese (60 U/ml) and 5 mmol/1 tliiosulphate protected islet cells from the toxic effect of 2.5 mmol/1 cyanide but not of 0.5 nmaol/1NP (Figs.…”

Section: Nitrite and Citrulline Production Correlates With Macrophagementioning

confidence: 97%

Cytotoxicity of activated rat macrophages against syngeneic islet cells is arginine-dependent, correlates with citrulline and nitrite concentrations and is identical to lysis by the nitric oxide donor nitroprusside

et al. 1993

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Lysis of rat islet cells by syngeneic activated macrophages in vitro can be completely inhibited by the nitric oxide-synthase-inhibitor NG-methyl-L-arginine. This inhibition can be reversed by an excess of L-arginine. Time-dependent lysis of islet cells by activated macrophages is accompanied by increasing concentrations of nitrite and citrulline in the culture medium both of which are measures of nitric oxide formation derived from L-arginine. Lysis of isolated islet cells and disintegration of isolated whole islets is also obtained within 15 h by culture in the presence of the nitric oxide generating vasodilator sodium nitroprusside. We thus conclude that nitric oxide is extremely toxic for islet cells and that nitric oxide alone and in the absence of other macrophage-generated potentially toxic products can rapidly and completely kill islet cells.

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“…It catalyzes the transfer of a sulfane sulfur atom from an anionic donor substrate to a thiophilic acceptor by means of a sulfur-substituted enzyme, covalent intermediate. This intermediate enzyme-sulfur complex was demonstrated to be a persulfide group (Cys-SSH) that can stably exist in neutral solution (22,24).…”

Section: Discussionmentioning

confidence: 99%

Formation of a selenium-substituted rhodanese by reaction with selenite and glutathione: Possible role of a protein perselenide in a selenium delivery system

Ogasawara

Lacourciere²,

Stadtman³

2001

Proc. Natl. Acad. Sci. U.S.A.

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Selenophosphate is the active selenium-donor compound required by bacteria and mammals for the specific synthesis of Secys-tRNA, the precursor of selenocysteine in selenoenzymes. Although free selenide can be used in vitro for the synthesis of selenophosphate, the actual physiological selenium substrate has not been identified. Rhodanese (EC 2.3.1.1) normally occurs as a persulfide of a critical cysteine residue and is believed to function as a sulfurdelivery protein. Also, it has been demonstrated that a seleniumsubstituted rhodanese (E-Se form) can exist in vitro. In this study, we have prepared and characterized an E-Se rhodanese. Persulfidefree bovine-liver rhodanese (E form) did not react with SeO 3 2؊ directly, but in the presence of reduced glutathione (GSH) and SeO 3 2؊ E-Se rhodanese was generated. These results indicate that the intermediates produced from the reaction of GSH with SeO 3 2؊ are required for the formation of a selenium-substituted rhodanese. E-Se rhodanese was stable in the presence of excess GSH at neutral pH at 37°C. E-Se rhodanese could effectively replace the high concentrations of selenide normally used in the selenophosphate synthetase in vitro assay in which the selenium-dependent hydrolysis of ATP is measured. These results show that a seleniumbound rhodanese could be used as the selenium donor in the in vitro selenophosphate synthetase assay.

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The differential functional stability of various forms of bovine liver rhodanese

Cited by 13 publications

References 30 publications

Mutagenic analysis of Thr‐232 in rhodanese from Azotobacter vinelandii highlighted the differences of this prokaryotic enzyme from the known sulfurtransferases

Mutagenic analysis of Thr‐232 in rhodanese from Azotobacter vinelandii highlighted the differences of this prokaryotic enzyme from the known sulfurtransferases

Cytotoxicity of activated rat macrophages against syngeneic islet cells is arginine-dependent, correlates with citrulline and nitrite concentrations and is identical to lysis by the nitric oxide donor nitroprusside

Formation of a selenium-substituted rhodanese by reaction with selenite and glutathione: Possible role of a protein perselenide in a selenium delivery system

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